Recent Advancements in Gene Expression and Enabling Technologies in Crop Plants

Kasi Azhakanandam • Aron Silverstone Henry Daniell • Michael R. DaveyEditors

Foreword by Mary-Dell Chilton, PhD, Syngenta Biotechnology, Inc.

Recent Advancements in Gene Expression and Enabling Technologies in Crop Plants

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ISBN 978-1-4939-2201-7 ISBN 978-1-4939-2202-4 (eBook)DOI 10.1007/978-1-4939-2202-4

Library of Congress Control Number: 2015931208

Springer New York Heidelberg Dordrecht London© Springer Science+Business Media, LLC 2015This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed.The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use.The publisher, the authors and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, express or implied, with respect to the material contained herein or for any errors or omissions that may have been made.

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Springer is part of Springer Science+Business Media (www.springer.com)

EditorsDr. Kasi AzhakanandamSyngenta Biotechnology Inc.3054 Cornwallis RdResearch Triangle Park, NC 27709, USA

Dr. Aron SilverstoneSyngenta Biotechnology Inc.3054 Cornwallis RdResearch Triangle Park, NC 27709, USA

Dr. Henry DaniellUniversity of PennsylvaniaSchool of Medicine240 South 40th StreetPhiladelphia PA 19104, USA

Dr. Michael R. DaveyUniversity of NottinghamSchool of BiosciencesLoughborough LE12 5RD,United Kingdom

This book is dedicated to people who have died of starvation

vii

Foreword

In the following pages, some of the world’s most renowned researchers take a look at the state of the art and science of introducing novel genes into plant cells and plants. The various chapters deal with a wide range of products, from genetically modified seeds and plants to commodities made by such transgenic plants, includ-ing enzymes or vaccines. One important consideration is where and how the new genes are integrated into the host plant. The donor DNA may be inserted into the plant chromosome at random places or targeted to a specific location, by recombi-nation or by employing site-specific nucleases. A future targeting technology may employ a minichromosome, an artificial vector assembled from parts of a normal chromosome (Chapter 13). A minichromosome is actually a megavector, which will be especially attractive for the introduction of a block of genes, for example those encoding an entire biochemical pathway for production of a valuable metabolite. At the other extreme of size, free replicons such as a (modified) plant DNA viral genome might be the most useful vector for some traits. Whatever the form and location of the vector, the DNA construct itself must mimic the plant’s strategy for dictating quantity, timing, and location for the encoded protein to be made. In Chapter 2, Dr. Nuccio et al. provides a wellspring of information on plant trait gene design and approaches that have worked.

This book addresses many of these issues and will be useful to the plant genetic engineer, whether student or accomplished professional. I found new ideas and information in each chapter. I skipped around as my curiosity led me, and was excited to discover how many different types of challenges plant genetic engineer-ing has posed, and how many creative solutions have been devised. I found the book quite readable for a technical work, with a refreshing honesty about the sometimes halting progress of scientific research.

While we are on the topic of honesty, I must confess to a motive underlying my writing of this foreword. I wanted to reach you, readers of this book, with one more message. Let me begin with a brief story: When my sons were quite young, we subscribed to a journal about the environment called Ranger Rick. One month it carried a story about insect galls, describing how the mother insect uses chemical signals to stimulate growth of the plant cells into a gall at the site where she deposits her eggs. When the insect larvae hatch, the gall serves her babies as a nice source

viii Foreword

of food. By coincidence, my colleagues and I at the University of Washington had recently begun a research project on crown gall tumors, induced by Agrobacterium in plants. The insect gall story, aimed at children, made me think. Crown galls were known to produce new metabolites—octopine or nopaline, depending on the Agro-bacterium strain that incited the gall. Could octopine and nopaline be baby food for Agrobacterium? When it was my turn to talk at our weekly research group meeting, I reported on the Ranger Rick article, and proposed that Agrobacterium, like the mother insect, might be producing the crown gall as a means of feeding its progeny. I can well recall the laughter and ridicule that ensued. The concept was named the Ranger Rick Hypothesis, and I was teased mercilessly about it for many months, until our competitors in France, Australia, and Belgium announced this very same concept as the “rationale of the gall” (in three languages). It became a respectable idea, eventually supported by increasing amounts of evidence.

There are several potential morals to this story, and I invite you to consider any of them that interest you. For me, the moral is that Agrobacterium truly was a genet-ic engineer before my colleagues and I ever thought of the possibility. The process that we now use to make genetically modified plants, the topic of this volume, is a natural one at core, invented first by a microbe and only refined by Homo sapi-ens. Agrobacterium worked out a way to transfer its desirable genes to the host plant cells, genes that caused abundant growth (the gall) and delicious (we suppose) meals for future generations. I hope that you who take a serious interest in the con-tents of this book will take equally seriously the need to inform the public that gene transfer is a natural and normal process. The products made by genetic modification of plants are more precise and predictable than those made by plant breeding, es-pecially plant breeders use of wide crosses for introduction of new traits from wild relatives of crop plants.

By the year 2050, the world’s population is expected to grow from its current 7 billion to 9 billion, a 30 % increase in the number of people. A distressing number of our present population is already hungry, even starving. Biotechnology alone cannot solve this problem, but it certainly has the potential to be an important part of the solution. Unless people accept foods produced through biotechnology, progress in food security will be slow. I believe that the principal risk of genetically modi-fied crops is public perception, not the safety of the products themselves, which are thoroughly tested. If you share my view, I hope that you will not keep it a secret. Seek opportunities to speak to school children, garden clubs, church groups, or anyone who will listen. Tell them that there is nothing unnatural about gene transfer to plants by Agrobacterium. I believe that the success of genetically modified plant products depends upon the efforts of scientists like you and me to communicate to the public the safety and sanity of biotech plants.

Mary-Dell ChiltonResearch Triangle Park, NC, USA

ix

Preface

When we decided to edit a book on gene expression in plants, we realized that the most valuable contribution would be to combine reports from the biotech industry, and academic and research institutes that would focus on gene expression studies with economically important crops and related enabling technologies. Such a vol-ume should be useful for students and researchers at all levels. Tremendous prog-ress has been made in introducing novel genes and traits into plant genomes since the first creation of transgenic plants 30 years ago, and the first commercialization of genetically modified maize in 1996. Consequently, cultivation of biotech crops with useful traits has increased more than 100-fold from 1.7 million ha in 1996 to over 175 million ha globally in 2013. This achievement has been made possible by continued advances in understanding the basic molecular biology of regulatory sequences to modulate gene expression, enhancement of protein synthesis, and new technologies for transformation of crop plants.

In this book, authors who are experts in their fields describe current advances on commercial crops and key enabling technologies that will underpin future ad-vances in biotechnology. They discuss state-of-the-art discoveries as well as future challenges. This book has three parts that encompass knowledge on genetically modified (GM) food crops that are currently used by consumers, those that are anticipated to reach the market place in the near future and enabling technologies that will facilitate the development of next generation GM crops. Part I focuses only on genetically modified maize and soybean (three chapters each), while Part II dis-cusses the GM food crops rice, wheat, sorghum, vegetables, and sugarcane. Part III covers exciting recent developments in several novel enabling technologies, includ-ing gene targeting, minichromosomes, and in planta transient expression systems.

In the first chapter, Lu et al. provide a detailed overview of fascinating aspects of maize protein expression. This chapter reviews current understanding and fu-ture perspectives on key aspects that affect recombinant protein expression in this crop. These authors have summarized various factors that control gene expression, including promoters, subcellular targeting, and different regulatory elements, in-cluding introns, 5ʹ and 3ʹ untranslated regions (UTRs), spacers and insulators. In Chapter 2, Nuccio et al. present a detailed understanding on transgene design with

x Preface

plant trait gene expression cassette design. The authors characterized several native maize promoters, and used the structure of these promoters to design constructs that deliver high-level gene expression/accumulation in maize. Chapter 3 is also devot-ed to maize. Howard and Hood review different strategies to maximize recombinant protein expression in kernels and discuss the characteristics that make maize a pop-ular choice for recombinant protein production. These authors also assess various factors that contribute to high-level expression of heterologous proteins, together with examples of successful approaches.

In Chapter 4, Ramachandra et al. outline the breeding and biotech approaches to improve yield in soybean. The use of transgenes to complement traditional breed-ing through “gene stacking” will be important to further increase soybean yield and overcome biotic and abiotic stresses. One of the most successful innovations of biotech that had a major impact on farming is the introduction of herbicide toler-ance in plants. Consequently, Huang et al. in Chapter 5 discuss the details of genes/traits, which have been exploited to make plants tolerant to herbicides. Tolerance to broad-spectrum herbicides makes weed control more efficient, which greatly assists the farming community. However, the increase of resistant weeds is creating new challenges for the biotech industry. In order to address this concern, authors discuss the use of trait stacking to manage hard-to-control and resistant weeds. They also describe the development of a new herbicide trait system for dicamba tolerance. Herman and Schmidt (Chapter 6) have focused on modification of soybean seeds for their use as protein bioreactors. Soybean seeds have high protein content and are used as a protein source in animal feed. These authors present the success and limitations of different approaches to produce heterologous proteins in seeds. They describe a protein rebalancing approach that increases expression of a model pro-tein (green fluorescent protein) from 1.5 to 8 % of the total soy seed protein.

Significant progress has been made in cereal biotechnology. Many traits have been engineered into the rice genome to protect against biotic and abiotic stress or to improve grain and nutritional quality. In Chapter 7, Nandi and Khush review strategies to increase heterologous protein expression in rice grains. These authors summarize key factors responsible for controlling expression, including regulatory sequences, translational efficiency, posttranslational modifications, and compart-mentalization of foreign proteins. They also discuss strategies to down-regulate en-dogenous protein expression in order to boost heterologous protein accumulation. In Chapter 8, Jones summarizes current advances in wheat biotechnology, particularly methods adopted for wheat transformation. He also summarizes progress in enhanc-ing tolerance to biotic stress and to improve quality traits such as those for bread-making. Biotechnology plays an important role in meeting the global demand for wheat, which is anticipated to increase more than 50 % by 2050. Recent advances in sorghum biotechnology are outlined by Do and Zhang (Chapter 9), with the chal-lenges related to the tissue culture and transformation of this crop. The biotech ap-proaches for insect pest management in vegetable crops are featured in Chapter 10 by Sreevathsa et al. The Bt protein was tested in vegetable crops to control insect pests, with discussion of different promoters used to achieve high-level expression, conferring greater resistance against target pests. The authors also discuss other

xiPreface

strategies, including the use of inhibitors of insect digestive enzymes, or engineer-ing secondary metabolism of volatile communication compounds to combat pests. In recent years, there has been more biotechnology research directed to sugarcane not only for sugar production, but also for its use as biofuels. In Chapter 11, Wu dis-cusses techniques for boosting sugar content through genetic engineering, including the expression of novel sugars.

As the opportunities of biotechnology increase, more complex tools are needed to deliver desired targets. In addition, newly acquired plant genomes’ sequences provide a wealth of data that can be exploited. A key to understanding the functions of specific genes is the ability to rapidly overexpress or turn them off. Part III ex-plores these enabling technologies. In Chapter 12, Petolino et al. describe gene tar-geting in plants by using Zinc Finger Nucleases (ZFNs). These authors explain how ZFNs are exploited for target mutagenesis, gene deletion and site-specific transgene integration. They also discuss other nuclease technologies, such as TALENs, mega-nucleases, and CRISPRs, as well as the relative advantages and limitations of these procedures. Minichromosomes combine native chromosome structural elements, like centromeres, along with transgenes for introduction into crop plants. Birchler (Chapter 13) reviews the status of “Minichromosome” technology in plants. One of the key advantages of artificial chromosomes is that multiple genes of interest could be stacked into plant genomes as a single entity without linkage to other chro-mosomes. Birchler also discusses both the challenges and opportunities associated with this novel technology.

Studies on gene function(s) utilizing stable transformation is time consuming and expensive. However, in planta transient sytems, using viral vectors developed in recent years, make it possible to study gene function by knocking down target genes or overexpression of genes of interest, although this approach has been lim-ited to small genes (< 1.5 kb) in crop plants. There are efforts to build viral vectors, which can accommodate larger inserts. In Chapter 14, Lee et al. review various in planta transient expression systems for both RNAi-mediated down-regulation and over expression of target genes in monocotyledonous plants. These authors discuss the increasing use of transient in planta expression systems, such as virus-induced gene silencing (VIGS), virus-mediated overexpression (VOX), and cell culture-based transient approaches, as well as the advantages and disadvantages associated with each transient system. Chapter 15 by Whitham et al. presents recombinant plant viruses that are capable of carrying genetic payloads of whole genes or gene fragments that provide convenient platforms as vectors for transient gene expres-sion and silencing in soybean. These authors focus on seven viral vector systems that have been used in this leguminous crop for VOX and/or VIGS applications. They discuss key features of the viral genomes, and future prospects to exploit viral vectors for soybean improvement.

In summary, this volume highlights a wide range of research tools, current methods, and future enabling technologies to improve crop plants to meet the ever increasing global demand for food, feed, and fuel. The editors believe that this book will be an excellent reference source for the scientific community interested in ex-tending model plant systems into valuable applications in crop plants. We sincerely

xii Preface

thank all the authors for their hard work and valuable contributions, and colleagues at Springer for the invitation to edit this unique contribution to the literature for the scientific community.

Kasi AzhakanandamAron Silverstone

Henry DaniellMichael R. Davey

xiii

Abstract

In the past two decades, agricultural biotechnology has had a major impact on farm-ing, with genetically modified (GM) crops grown on more than 175 million ha glob-ally. Although plant biotechnology has exploited model systems to gain fundamen-tal knowledge, parallel research on field-grown plants has facilitated the develop-ment of GM crops that are used by consumers today. Biotechnology has also helped to create a rich pipeline of future products. This volume focuses on the innovations in both applied and basic research that are advancing our ability to deliver more complex multigene traits into plants. Although much of the work to date has been done on corn and soybean, other plants that are the subject of active transgenic de-velopment include rice, wheat, sorghum, sugarcane, and vegetable crops. There is a progression from the use of constitutive promoters and single traits to gene stack-ing, the design of transgene cassettes to more resemble native genes, the subcellular location of recombinant proteins, and manipulating storage tissues to achieve op-timal performance. Herbicide tolerance and insect control have been and will con-tinue to be highly desired traits. The future holds promise for novel modes of action to overcome current limitations. Targets for engineered recombinant proteins go beyond agronomic traits and focus on industrial or pharmaceutical uses, yield, and nutritional enhancement. Undoubtedly, future farming will advance from food/feed to industrial products, making crops more rewarding with value-added traits. Soon, even more sophisticated tools, including precision insertion or editing of genes and building novel chromosomes, will increase our ability to overcome current barriers in gene expression technology and facilitate rapid regulatory approval. The use of transient expression systems for crop plants will facilitate rapid evaluation of trans-genes in crop plants. This book highlights a wide range of current research tools and enabling technologies to improve crop plants, with special emphasis on next generation approaches for engineering complex traits and value-added products that will revolutionize the future of agriculture to meet the ever increasing global demand for food, feed, fuel, and industrial products.

xv

Acknowledgements

The editors would like to thank Rongda Qu, Todd J. Jones, Michael Nuccio, Albert Lu, Bernard Vernooij, Jared Conville, Sandeep Kumar, Robert Thomas Gaeta, P. Dayanandan, Brian Hague, and Vidya Rajan for reviewing several of the chapters.

xvii

Part I Corn and Soybean

1 Maize Protein Expression ���������������������������������������������������������������������������� 3Albert Lu, Scott Diehn and Mark Cigan

2 Plant Trait Gene Expression Cassette Design ����������������������������������������� 41Michael Nuccio, Xi Chen, Jared Conville, Ailing Zhou and Xiaomei Liu

3 Strategies to Maximize Recombinant Protein Expression in Maize Kernels ���������������������������������������������������������������������������������������� 79John A� Howard and Elizabeth E� Hood

4 Breeding and Biotech Approaches Towards Improving Yield in Soybean ��������������������������������������������������������������������������������������������������� 131Dhanalakshmi Ramachandra, Savitha Madappa, Jonathan Phillips, Paul Loida and Balasulojini Karunanandaa

5 Towards Using Biotechnology to Modify Soybean Seeds as Protein Bioreactors ������������������������������������������������������������������������������������ 193Eliot M� Herman and Monica A� Schmidt

6 Herbicide Tolerance ����������������������������������������������������������������������������������� 213Jintai Huang, Christine Ellis, Brian Hauge, Youlin Qi and Marguerite J� Varagona

Part II Other Economically Important Crops

7 Strategies to Increase Heterologous Protein Expression in Rice Grains ��������������������������������������������������������������������������������������������� 241Somen Nandi and Gurdev S� Khush

Contents

xviii Contents

8 Wheat Biotechnology: Current Status and Future Prospects������������ 263Huw D� Jones

9 Sorghum Transformation: Achievements, Challenges, and Perspectives ������������������������������������������������������������������������������������� 291Phat T� Do and Zhanyuan J� Zhang

10 Biotechnology for Insect Pest Management in Vegetable Crops ������� 313Rohini Sreevathsa, Amolkumar U� Solanke and P� Solanke and P� Solanke and P Ananda Kumar

11 Enhancement of Sugar Yield by Introducing a Metabolic Sink in Sugarcane����������������������������������������������������������������������������������� 341Luguang Wu

Part III Enabling Technologies

12 Zinc Finger Nuclease-Mediated Gene Targeting in Plants����������������� 363Joseph F� Petolino, Lakshmi Sastry-Dent and J� Pon Samuel

13 Engineered Minichromosome Technology in Plants��������������������������� 383James A� Birchler

14 In Planta In Planta Transient Expression Systems for Monocots ���������������������� 391Wing-Sham Lee, Kim E� Hammond-Kosack and Kostya Kanyuka

15 Recent Advances in In Planta Transient Expression and Silencing Systems for Soybean Using Viral Vectors���������������������������� 423

Steven A� Whitham, Alan L� Eggenberger, Chunquan Zhang, R� V� V� V Chowda-Reddy, Kathleen M� Martin and John H� Hill

Erratum ���������������������������������������������������������������������������������������������������������� E1

Erratum ���������������������������������������������������������������������������������������������������������� E3

Index���������������������������������������������������������������������������������������������������������������� 453

xix

Contributors

James A. Birchler Division of Biological Sciences, University of Missouri, Columbia, MO, USA

Xi Chen Syngenta Biotechnology, Inc� East Cornwallis Road, Research Triangle Park, NC, USA

R. V. Chowda-Reddy Department of Plant Pathology and Microbiology, Iowa State University, Ames, IA, USA

Mark Cigan DuPont Pioneer, Johnston, IA, USA

Jared Conville Syngenta Biotechnology, Inc�, Research Triangle Park, NC, USA

Scott Diehn DuPont Pioneer, Johnston, IA, USA

Phat T. Do Plant Transformation Core Facility, Division of Plant Sciences, 007A, Sears Plant Growth Facility, University of Missouri, Columbia, MO, USA

Alan L. Eggenberger Department of Plant Pathology and Microbiology, Iowa State University, Ames, IA, USA

Christine Ellis Monsanto Company, Chesterfield, MO, USA

Kim E. Hammond-Kosack Wheat Pathogenomics Team, Plant Biology and Crop Science Department, Rothamsted Research, Harpenden, UK

Brian Hauge Monsanto Company, Chesterfield, MO, USA

Eliot M. Herman School of Plant Sciences, BIO5 Institute Room No� 249, University of Arizona, Tucson, AZ, USA

John H. Hill Department of Plant Pathology and Microbiology, Iowa State University, Ames, IA, USA

Elizabeth E. Hood Biosciences Institute, Arkansas State University, State University, AR, USA

John A. Howard Cal Poly Technology Park, San Luis Obispo, CA, USA

xx Contributors

Jintai Huang Monsanto Company, Chesterfield, MO, USA

Jones Huw D. Rothamsted Research, Harpenden, Hertfordshire, UK

Kostya Kanyuka Wheat Pathogenomics Team, Plant Biology and Crop Science Department, Rothamsted Research, Harpenden, UK

Balasulojini Karunanandaa Monsanto Company, Chesterfield, MO, USA

Gurdev S. Khush Global HealthShare® Initiative, Department of Plant Science, University of California, Davis, CA, USA

P. Ananda Kumar Institute of Biotechnology, ANGRAU, Hyderabad, India

Wing-Sham Lee Wheat Pathogenomics Team, Plant Biology and Crop Science Department, Rothamsted Research, Harpenden, UK

Xiaomei Liu Syngenta Biotechnology, Inc�, Research Triangle Park, NC, USA

Paul Loida Monsanto Company, Chesterfield, MO, USA

Albert Lu DuPont Pioneer, Johnston, IA, USA

Savitha Madappa Monsanto Company, Bangalore, India

Kathleen M. Martin United States Department of Agriculture, Agricultural Research Service, Corn Insects and Crop Genetics Research Unit, Ames, IA, USA

Somen Nandi Global HealthShare® Initiative, Department of Molecular and Cellular Biology, University of California, Davis, CA, USA

Michael Nuccio Syngenta Biotechnology, Inc�, Research Triangle Park, NC, USA

Joseph F. Petolino Dow AgroSciences, Indianapolis, IN, USA

Jonathan Phillips Monsanto Company, St� Louis, MO, USA

Youlin Qi Monsanto Company, Chesterfield, MO, USA

Dhanalakshmi Ramachandra Monsanto Company, Bangalore, India

J. Pon Samuel Dow AgroSciences, Indianapolis, IN, USA

Lakshmi Sastry-Dent Dow AgroSciences, Indianapolis, IN, USA

Monica A. Schmidt School of Plant Sciences, BIO5 Institute Room No� 303, University of Arizona, Tucson, AZ, USA

Amolkumar U. Solanke National Research Centre for Plant Biotechnology, New Delhi, India

Rohini Sreevathsa National Research Centre for Plant Biotechnology, New Delhi, India

Marguerite J. Varagona Biotechnology-Agronomic Traits, Monsanto Company, Chesterfield, MO, USA

xxiContributors

Steven A. Whitham Department of Plant Pathology and Microbiology, Iowa State University, Ames, IA, USA

Luguang Wu School of Agriculture and Food Science, Faculty of Science, The University of Queensland, St� Lucia, Qld, Australia

Chunquan Zhang Department of Agriculture, Alcorn State University, Lorman, MS, USA

Zhanyuan J. Zhang Plant Transformation Core Facility, Division of Plant Sciences, 007A, Sears Plant Growth Facility, University of Missouri, Columbia, MO, USA

Ailing Zhou Syngenta Biotechnology, Inc�, Research Triangle Park, NC, USA

xxiii

About the Editors

Dr. Kasi Azhakanandam earned his Bachelor, Master, and MPhil degrees in Bi-ology from Madras Christian College, the University of Madras, India and a PhD in Plant Biotechnology from the University of Nottingham, UK. He worked as a Guest Lecturer at Madras Christian College for a short period before joining Mahyco, India, as a Deputy Chief Scientist/Principal Investigator, where he established a crop transformation laboratory. He led a team, which established transformation in commercial Indica rice, Indian cotton varieties, and six different vegetable crops, including Bt eggplant; these are waiting for approval for commercial cultivation in India while the Bt eggplant is approved for commercial cultivation in Bangladesh. He also successfully produced marker-free rice and vegetable crops. Following his postdoctoral work related to vaccine production for cervical cancer at North Caro-lina State University, Dr. Azhakanandam joined Syngenta Biotechnology, Inc., at Research Triangle Park, NC as a Staff Scientist III. He has worked on a range of projects to improve crops through genetic engineering, and currently leads a techni-cal team for developing new traits for corn.

Dr. Aron Silverstone gained a Bachelor’s degree in Biology from Harvard Uni-versity, and a PhD in Plant Physiology from the University of California, Davis. He conducted his postdoctoral research at Duke University’s Department of Botany, studying gibberellin biosynthesis and response. Following his postdoctoral work, Dr. Silverstone joined Syngenta Biotechnology, Inc., at Research Triangle Park, NC as a Staff Scientist I. He has worked on several projects in corn, soy and sugarcane to improve crops through genetic engineering. Dr. Silverstone is currently working on protecting plants from abiotic stresses.

Dr. Henry Daniell received his education in India, and is currently a Professor and Director of Translational Research at the University of Pennsylvania. He is a Fellow of the American Association for the Advancement of Science and a for-eign member of the Italian National Academy of Sciences (14th American to be inducted in the past 230 years). He is the editor-in-chief of the Plant Biotechnology Journal, Oxford, UK. Dr. Daniell is the recipient of several awards, including the American Diabetes Association Award, Bayer Hemophilia Global Award, and Bill

xxiv About the Editors

and Melinda Gates Foundation Award, for his scientific contributions. He is recog-nized for pioneering chloroplast genetic engineering as a new platform to produce and deliver orally low-cost vaccines and biopharmaceuticals bioencapsulated in plant cells. His invention was ranked by Nature Biotechnology among the top ten inventions of the past decade and among Biomed Central’s Hot 100 authors in the world. He has more than 150 published patents and over 200 scientific publications.

Dr. Michael R. Davey has a BSc Honours degree in Botany from University Col-lege, Swansea, Wales, and a PhD from the University of Leicester, UK. In 1970, he was appointed to a research position at the University of Nottingham, UK where he continued his work on plant ultrastructure and gene transfer techniques. He has published extensively on plant cell culture and genetic engineering, and holds an Honorary Lectureship in the School of Biosciences, University of Nottingham.